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BMC Hematology

, 16:9

Hemostasis, thrombosis and vascular biology

Abstract

BackgroundFibrin provides a temporary matrix at the site of vascular injury. The aims of the present work were 1 to follow fibrin formation and lysis onto the surface of human dermal microvascular endothelial cells HMEC-1, and 2 to quantify the secretion of fibrinolytic components in the presence of fibrin.

MethodsFibrin clots at different fibrinogen concentrations were formed on top of model 1 or beneath model 2 the endothelial cells. Fibrin formation or lysis onto the surface of HMEC-1 cells, was followed by turbidity. Clot structure was visualized by laser scanning confocal microscopy LSCM. The secretion of uPA and PAI-1 by HMEC-1 cells was quantified by ELISA.

ResultsThe rate of fibrin formation increased approximately 1.5-fold at low fibrinogen content 0.5 and 1 mg-mL; p < 0.05 compared to the condition without cells; however, it was decreased at 2 mg-mL fibrinogen p < 0.05 and no differences were found at higher fibrinogen concentrations 3 and 5 mg-mL. HMEC-1 retarded dissolution of clots formed onto their surface at 0.5 to 3 mg-mL fibrinogen p < 0.05. Secretion of uPA was 13 × 10 ng-mL per cell in the absence of RGD and 8 × 10 ng-mL per cell in the presence of RGD, when clots were formed on the top of HMEC-1. However, the opposite was found when cells were grown over fibrin: 6 × 10 ng-mL per cell without RGD vs. 17 × 10 ng-mL per cell with RGD. The secretion of PAI-1 by HMEC-1 cells was unrelated to the presence of fibrin or RGD, 7 × 10 μg-mL per cell and 5 × 10 μg-mL per cell, for the apical model 1 and basal clots model 2, respectively.

ConclusionsHMEC-1 cells influence fibrin formation and dissolution as a function of the fibrin content of clots. Clot degradation was accentuated at high fibrin concentrations. The secretion of fibrinolytic components by HMEC-1 cells seemed to be modulated by integrins that bind RGD ligands.

KeywordsFibrinogen Fibrin Fibrinolysis Urokinase-type plasminogen activator Plasminogen activator inhibitor type 1 Endothelium Human dermal microvascular endothelial cells AbbreviationsECsendothelial cells

ELISAenzyme-linked immunosorbent assay

Fgfibrinogen

FXIIIaactivated factor XIII

HMEChuman dermal microvascular endothelial cells

HOMChuman mesothelial cells

HOTMChuman omental tissue microvascular endothelial cells

HUVEChuman umbilical vein endothelial cell

LRlysis rate

LSCMlaser scanning confocal microscopy

MaxAbsmaximum absorbance

ODoptical density

PAI-1plasminogen activator inhibitor type 1

PBSphosphate buffered-saline

Pgplasminogen

RGDarginyl glycyl aspartic acid

SEMstandard error of the mean

tPAtissue type plasminogen activator

uPAurokinase-type plasminogen activator

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Autor: O. Castillo - H. Rojas - Z. Domínguez - E. Anglés-Cano - R. Marchi

Fuente: https://link.springer.com/article/10.1186/s12878-016-0048-6







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